The invention described herein relates to diagnostic and therapeutic methods targeting cell signaling pathways perturbed in syndromes involving disregulated cellular proliferation.
Cancers are the second most prevalent cause of death in the United States, causing 450,000 deaths per year. One in three Americans will develop cancer, and one in five will die of cancer. While substantial progress has been made in identifying some of the likely environmental and hereditary causes of cancer, there is a need for additional diagnostic and therapeutic modalities that target cancer and related diseases and disorders.
A number of so-called cancer genes, i.e., genes that have been implicated in the etiology of cancer, have been identified in connection with hereditary forms of cancer and in a large number of well-studied tumor cells. Cancer genes ate broadly classified into xe2x80x9concogenesxe2x80x9d which, when activated, promote tumorigenesis, and xe2x80x9ctumor suppressor genesxe2x80x9d which, when damaged, fail to suppress tumorigenesis. While these classifications provide a useful method for conceptualizing tumorigenesis, it is also possible that a particular gene may play differing roles depending upon the particular allelic form of that gene, its regulatory elements, the genetic background and the tissue environment in which it is operating.
The HER-2/neu is an illustrative oncogene that is member of the tyrosine protein kinase family of oncogenes and shares a high degree of homology with the epidermal growth factor receptor and presumably plays a role in cell growth and/or differentiation. Amplification and/or overexpression of the HER-2/neu type I receptor tyrosine kinase has been detected in 25-30% of human breast cancers. This alteration is an independent prognostic factor predicting poor clinical outcome. HER-2/neu overexpression in experimental models has been associated with an aggressive phenotype, including increased proliferation rate, DNA synthesis, anchorage-independent growth, tumorigenicity and metastatic potential. Recently, this genetic alteration has been successfully targeted in humans using the monoclonal antibody HERCEPTIN. Despite these advances, the molecular pathways leading from HER-2/neu overexpression to increased malignancy remain unclear and few targets downstream of HER-2/neu have been identified. Much remains to be learned about how HERCEPTIN functions as an anti-tumor agent and why certain HER-2/neu overexposing tumors respond to therapy while many others do not.
Ligands of the TGF-xcex2 receptor (TGF-xcex21, TGF-xcex22 and TGF-xcex23) are also known to play a role in syndromes involving the disregulation of cellular growth. Specifically, TGF-xcex2 ligands are involved in the growth inhibition in a number of cell types, and loss of this negative regulation is thought to contribute to tumor development. Studies have demonstrated that TGF-xcex2 ligands suppress the growth of certain cancer cell lines, that antisense inhibition of TGF-xcex2 enhances the tumorigenicity of weakly tumorigenic cancer cell lines, and that certain tumor cells are unresponsive to growth inhibition by TGF-xcex2 ligands.
While researchers have identified a variety of genes involved in growth disregulation such as Her-2/neu and TGF-xcex2, there is need for more detailed analyses of the genetic profiles exhibited by cells in which the regulatory processes that control cell growth have been disrupted. Moreover, an understanding of how the products of genes involved in disregulated cell growth interact in a larger context is needed for the development of improved diagnostic and therapeutic methods for identifying and treating pathological syndromes associated with growth disregulation such as cancer.
The present invention provides methods for obtaining genetic profiles of cancer cells in order to assess the status of a cancer in an individual. In addition, the present invention provides methods for inhibiting the growth of cancer cells that exhibit certain genetic profiles. These methods identify an important link between HER-2/neu overexpression and loss of growth inhibition by the TGF-xcex2 signaling pathway in cancer cells.
The invention disclosed herein provides a number of embodiments pertaining to the analysis of gene expression of mammalian cells. In these methods, the mRNA expression of a plurality of genes is examined in order to gain information that is useful in the diagnosis and/or prognosis of a pathological condition such as cancer. A typical embodiment consists of a method of examining a breast cell for evidence of a malignant phenotype by examining the mRNA expression profile of at least two mRNA polynucleotides in the cell, wherein the mRNA polynucleotides are selected from a first group of mRNA polynucleotides that are overexpressed in cells that overexpress Her-2/neu and a second group of mRNA polynucleotides that are underexpressed in cells that overexpress Her-2/neu. An mRNA expression profile in which the mRNA levels of the mRNA polynucleotides in the first group are increased greater than about 2 fold relative to the corresponding mRNA levels of the mRNA polynucleotides in normal breast cells and the mRNA levels of the mRNA polynucleotides in the second group are decreased greater than about 2 fold relative to the corresponding mRNA levels of the mRNA polynucleotides in normal breast cells is indicative of a malignant phenotype.
In preferred embodiments of the invention, the mRNA expression profile of at least one mRNA polynucleotide from the first group is examined and the mRNA expression profile of at least one mRNA polynucleotide from the second group is examined. In highly preferred embodiments, the mRNA expression profile of at least 3, 4, 5, 6, 7, 8, 9, 10 or all of the mRNA polynucleotides is examined. While the mRNA expression profile of genes can be evaluated by a wide variety of methods known in the art, in preferred embodiments, the mRNA expression profile is examined using a cDNA microarray.
Another related embodiments of the invention that pertains to the analysis of gene expression consists of a method for examining a breast cell for evidence of a malignant phenotype by comparing the level of mRNA expression of bone morphogenetic protein 7 (BMP-7) with the level of mRNA of at least one comparative gene, wherein an mRNA expression profile in which levels of BMP-7 mRNA are lower than those of the comparative gene is indicative of a malignant phenotype. In preferred embodiments of the invention, the level of BMP-7 mRNA expression is compared to 2, 3, 4, 5, 6, 7 or 8 comparative genes is examined. In a further embodiment of the invention, the cytological characteristics of the cell are examined, preferably for mesenchymal characteristics and/or epithelial characteristics.
Building upon the data generated in the profiling experiments, subsequent experiments revealed that an antibody capable of inhibiting Her-2/neu receptor function and a TGF-xcex2 family member can act synergistically to inhibit the growth of mammalian cells, particularly mammalian cancer cells. The invention provides various methods that involve the use of an antibody capable of inhibiting Her-2/neu receptor function and a TGF-xcex2 family member for inhibiting the growth of mammalian cells. For example, the invention provides a method for inhibiting the growth of mammalian cells comprising exposing a mammalian cell, such as a cancer cell (preferably a breast or ovarian cancer cell), to an antibody capable of inhibiting Her-2/neu receptor function and a TGF-xcex2 family member in an amount effective to synergistically inhibit cell growth. The cell may be in cell culture or in a mammal, e.g. a mammal suffering from cancer or a condition in which inhibiting the growth of the cells is desirable. Thus, the invention includes a method for treating a mammal suffering from cancer comprising administering an effective amount of an antibody capable of inhibiting Her-2/neu receptor function and a TGF-xcex2 family member, as disclosed herein.
The invention also provides compositions which comprise an antibody capable of inhibiting Her-2/neu receptor function and/or a TGF-xcex2 family member. Optionally, the compositions of the invention will include pharmaceutically acceptable carriers or diluents. Preferably, the compositions will include an antibody capable of inhibiting Her-2/neu receptor function and/or a TGF-xcex2 family member in an amount which is effective to synergistically inhibit the growth of mammalian cells.
The invention also provides articles of manufacture and kits which include reagents for analyzing the expression of genes within a cell, an antibody capable of inhibiting Her-2/neu receptor function and/or a TGF-xcex2 family member.